This project focuses on the regulation and function of the MAPK Kinase Kinase MEKK3. MAPK pathways have been shown as essential to cell growth, differentiation and apoptosis. It is therefore of interest to gain a better molecular understanding of i) all of the component parts that make up these signaling paths, and ii) of the physiologic targets of individual MAPK pathways in normal and diseased states. We originally cloned MEKK3 and determined that it can activate all three major MAPKs, namely ERK, JNK and p38, albeit to different extents. We also discovered that MEKK3 can arrest growth. In this regard, an estrogen-activatable form of this kinase could block cells in the G1 phase of the cell cycle in permanently transfected cells treated with estrogen. This arrest is mediated in part by negative regulation of cyclin D1 via a p38-dependent mechanism. MEKK3 also reversed Ras-induced cell transformation and may serve to play an important role to arrest cell cycle progression during cellular stress or damage. Regarding regulation of MEKK3, we discovered a direct interaction with and phosphorylation by the AKT kinase, a major mediator of anti-apoptotic pathways. In addition, we have cloned a novel MEKK3 interacting protein that may be a physiologic regulator of MEKK3 function.